In order to diagnose disease or the conditions of disease, detection of a specific biomolecule (biomarker) contained in blood and the like has been noted.
Presently, in order to detect the biomolecule, immunoassay employing an antigen-antibody reaction is mainly employed. A chemical luminescence method employing an enzyme reaction is presently prevailing on account of high sensitivity. However, the use of the enzyme, which is a biosubstance, requires complicated operations such as severe temperature control and the like. More simple systems are desired.
As one of the methods, there is known a fluorescence immunoassay method employing a biosubstance labeling agent labeled in advance with a fluorescent substance. As the fluorescent substance, an organic dye or a quantum dot can be used. The fluorescence immunoassay method can be carried out more simply as compared with the chemical luminescence method. However, the fluorescence intensity of the fluorescent substance used is extremely low, resulting in poor detection sensitivity.
Recently, in order to diagnose disease in an extremely early stage, i.e., to carry out detection of an extremely small amount of a biomarker, a biosubstance labeling agent labeled with a fluoresce substance emitting fluorescence with high density is demanded. As one method for obtaining such a substance, there is a method which incorporates a fluorescent substance in one silica nanoparticle (see, for example, Patent Documents 1 and 2).
Patent Document 3 discloses that fluorescent dye-containing silica particles are coated with silica in order to increase the emission luminance. Patent Document 4 discloses that silica particles containing a fluorescent dye in the connecting pores are coated with silica.
The present inventors prepared fluorescent dye-containing silica particles coated with silica according to the method as disclosed in Patent Document 3, and carried out fluorescence immunoassay after one month's storage. As a result, a ratio (S/N ratio) of the fluorescence intensity S due to the labeling agent combined with a biomarker to be detected to the fluorescence intensity N due to the labeling agent in the absence of a biomarker to be detected proved to be small.
The present inventors also prepared a quantum dot-containing silica prepared according to a method disclosed in Patent document 2, and prepared a quantum dot-containing silica with a silica coating according to a method disclosed in Patent document 3, Both quantum dot-containing silicas showed the result similar to that described above.
The above result means that when the fluorescent substance-containing silica prepared according to a known technique is applied to extracorporeal diagnosis medicines, there is problem in long term storage stability. Further improvement is desired.
Prior Art Document
Patent Document
    Patent Document 1: WO 07/074722    Patent Document 2: Japanese Patent O.P.I. Publication No. 2003-321226    Patent Document 3: Japanese Unexamined Patent Application Publication (Translation of PCT Application) No. 2006-514708    Patent Document 4: Japanese Patent O.P.I. Publication No. 2009-196829